The administration of immunostimulatory oligonucleotide sequences (ISS-ODN, also known as CpG-ODN) inhibits experimental asthma. This protective effect is transient and lasts for a few weeks. We recently identified that ISS-ODN induces high levels of pulmonary indoleamine 2,3-dioxygenase (IDO), a key enzyme of tryptophan (Trp) metabolism. Our preliminary data suggest that IDO may protect against experimental asthma by inducing apoptosis of antigen (i.e., allergen) stimulated Th2 cells via Trp deprivation in an organ-specific fashion (i.e., lung specific). Based on our observation and aligned with the exploratory nature of this RFA--specifically the identification of mechanisms by which known biological agents induce and/or maintain immune tolerance--we have put together two specific aims (SA). Thus, in SA-1 we hypothesize that the induction of transient tolerance is due to the depletion of antigen-specific pulmonary Th2 cells and that homeostatic proliferation, which replenishes this population with time, accounts for the re-emergence of the asthmatic phenotype. Recent published data indicate that IDO expressing antigen presenting cells (APCs) mediate a regulatory role on T cell responses and support the development of T regulatory (Treg) cells. Thus, in SA-2 we hypothesize that under the conditions of repeated ISS administration to chronically OVA-challenged mice an antigen specific and a long-term tolerance is developed secondary to the generation of Treg cells and their recruitment into the lungs. We will validate these hypotheses in a series of experiments in vivo using (1) an adoptive transfer-based approach of in-vitro differentiated, OVA-specific, transgenic Th2 cells to wt mice and (2) OVA/alum sensitized mice. The results of the proposed studies will offer new insight into controlling immune tolerance by providing the molecular/biochemical basis for tolerance induction by ISS-ODN. Furthermore, these results will help to optimize the therapeutic effects of ISS-ODN in coming clinical trials in patients with allergic asthma.